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Caloric restriction reverses high-fat diet-induced endothelial dysfunction and vascular superoxide production in C57Bl/6 mice

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Abstract

Obesity is frequently associated with endothelial dysfunction. We hypothesized that high-fat feeding dysregulates the balance between endothelial derived nitric oxide and superoxide formation. Furthermore, we examined whether caloric restriction could reverse the detrimental vascular effects related to obesity. Male C57Bl/6 mice were fed with normal-fat diet (fat 17%) or high-fat diet (fat 60%) for 150 days. After establishment of obesity at day 100, a subgroup of obese mice were put on caloric restriction (CR) (70% of ad libitum energy intake) for an additional 50 days. At day 100, aortic rings from obese mice receiving high-fat diet showed impaired endothelium-dependent vasodilation in response to acetylcholine (ACh). Caloric restriction reversed high-fat diet-induced endothelial dysfunction. At day 150, impaired vasodilatory responses to ACh in obese mice without caloric restriction were markedly improved by preincubation with the tetrahydrobiopterin (BH4) precursor sepiapterin and l-arginine, a substrate for endothelial nitric oxide synthase (eNOS). Additionally, inhibition of vascular arginase by l-norvaline partially, and superoxide scavenging by Tiron completely, restored endothelial cell function. Obese mice showed increased vascular superoxide production, which was diminished by endothelial denudation, pretreated of the vascular rings with apocynin (an inhibitor of reduced nicotinamide adenine dinucleotide phosphate [NADPH] oxidase), oxypurinol (an inhibitor of xanthine oxidase), N G-nitro-l-arginine methyl ester (LNAME; an inhibitor of eNOS), or by adding the BH4 precursor sepiapterin. Caloric restriction markedly attenuated vascular superoxide production. In obese mice on CR, endothelial denudation increased superoxide formation whereas vascular superoxide production was unaffected by l-NAME. Western blot analysis revealed decreased phosphorylated eNOS (Ser1177)-to-total eNOS expression ratio in obese mice as compared to lean controls, whereas the phospho-eNOS/NOS ratio in obese mice on CR did not differ from the lean controls. In conclusion, the present study suggests that caloric restriction reverses obesityinduced endothelial dysfunction and vascular oxidative stress, and underscores the importance of uncoupled eNOS in the pathogenesis.

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References

  1. Lindgren CM, McCarthy MI (2008) Mechanisms of disease: genetic insights into the etiology of type 2 diabetes and obesity. Natl Clin Pract Endocrinol Metab 4(3):156–163

    Article  CAS  Google Scholar 

  2. Avogaro A, de Kreutzenberg SV (2005) Mechanisms of endothelial dysfunction in obesity. Clin Chim Acta 360(1–2):9–26

    Article  CAS  PubMed  Google Scholar 

  3. Van Gaal LF, Mertens IL, De Block CE (2006) Mechanisms linking obesity with cardiovascular disease. Nature 444(7121): 875–880

    Article  PubMed  Google Scholar 

  4. Heistad DH (2006) Oxidative stress and vascular disease. Arterioscler Thromb Vasc Biol 26:689–695

    Article  CAS  PubMed  Google Scholar 

  5. Pechánová O, Simko F (2007) The role of nitric oxide in the maintenance of vasoactive balance. Physiol Res 56suppl 2:S7–S16

    PubMed  Google Scholar 

  6. Stuehr D, Pou S, Rosen GM (2001) Oxygen reduction by nitricoxide synthases. J Biol Chem 276(18):14533–14536

    Article  CAS  PubMed  Google Scholar 

  7. Takaya T, Hirata K, Yamashita T, Shinohara M, Sasaki N, Inoue N, Yada T, Goto M, Fukatsu A, Hayashi T, Alp NJ, Channon KM, Yokoyama M, Kawashima S (2007) A specific role for eNOS-derived reactive oxygen species in atherosclerosis progression. Arterioscler Thromb Vasc Biol 27(7):1632–1637

    Article  CAS  PubMed  Google Scholar 

  8. Alp NJ, Mussa S, Khoo J, Cai S, Guzik T, Jefferson A Goh N, Rockett KA, Channon KM (2003) Tetrahydrobiopterin-dependent preservation of nitric oxide-mediated endothelial function in diabetes by targeted transgenic GTP-cyclohydrolase I overexpression. J Clin Invest 112(5):725–735

    CAS  PubMed  Google Scholar 

  9. Mitchell BM, Cook LG, Danchuk S, Puschett JB (2007) Uncoupled endothelial nitric oxide synthase and oxidative stress in a rat model of pregnancy-induced hypertension. Am J Hypertens 20(12):1297–1304

    Article  CAS  PubMed  Google Scholar 

  10. Parekh PI, Petro AE, Tiller JM, Feinglos MN, Surwit RS (1998) Reversal of diet-induced obesity and diabetes in C57BL/6J mice. Metabolism 47(9):1089–1096

    Article  CAS  PubMed  Google Scholar 

  11. Traupe T, D’Uscio LV, Muenter K, Morawietz H, Vetter W, Barton M (2002) Effects of obesity on endothelium-dependent reactivity during acute nitric oxide synthase inhibition: modulatory role of endothelin. Clin Sci (Lond) 103suppl 48:13S–15S

    CAS  Google Scholar 

  12. Barton M, Carmona R, Morawietz H, d’Uscio LV, Goettsch W, Hillen H Haudenschild CC, Krieger JE, Münter K, Lattmann T, Lüscher TF, Shaw S (2000) Obesity is associated with tissue-specific activation of renal angiotensin-converting enzyme in vivo: evidence for a regulatory role of endothelin. Hypertension 35(1 Pt 2):329–336

    CAS  PubMed  Google Scholar 

  13. Mundy AL, Haas E, Bhattacharya I, Widmer CC, Kretz M, Hofmann-Lehmann R (2007) Fat intake modifies vascular responsiveness and receptor expression of vasoconstrictors: implications for diet-induced obesity. Cardiovasc Res 73(2):368–375

    Article  CAS  PubMed  Google Scholar 

  14. Traupe T, Lang M, Goettsch W Münter K, Morawietz H, Vetter W (2002) Obesity increases prostanoid-mediated vasoconstriction and vascular thromboxane receptor gene expression. J Hypertens 20(11):2239–2245

    Article  CAS  PubMed  Google Scholar 

  15. Molnar J, Yu S, Mzhavia N, Pau C, Chereshnev I, Dansky HM (2005) Diabetes induces endothelial dysfunction but does not increase neointimal formation in high-fat diet fed C57BL/6J mice. Circ Res 96(11):1178–1184

    Article  CAS  PubMed  Google Scholar 

  16. Noronha BT, Li JM, Wheatcroft SB, Shah AM, Kearney MT (2005) Inducible nitric oxide synthase has divergent effects on vascular and metabolic function in obesity. Diabetes 54(4): 1082–1089

    Article  CAS  PubMed  Google Scholar 

  17. Ketonen J, Merasto S, Paakkari I, Mervaala EM (2005) High sodium intake increases vascular superoxide formation and promotes atherosclerosis in apolipoprotein E-deficient mice. Blood Press 14(6):373–382

    Article  CAS  PubMed  Google Scholar 

  18. Ketonen J. Mervaala EMA (2008) Effects of dietary sodium on ROS formation and endothelial dysfunction in LDL receptor deficient mice on high fat diet. Heart Vessels 23(6):420–429

    Article  PubMed  Google Scholar 

  19. Ahotupa M, Marniemi J, Lehtimäki T, Talvinen K, Raitakari OT, Vasankari T (1998) Baseline diene conjugation in LDL lipids as a direct measure of in vivo LDL oxidation. Clin Biochem 31:245–261

    Article  Google Scholar 

  20. Furukawa S, Fujita T, Shimabukuro M, Iwaki M, Yamada Y, Nakajima Y, Nakayama O, Makishima M, Matsuda M, Shimomura I (2004) Increased oxidative stress in obesity and its impact on metabolic syndrome. J Clin Invest 114(12):1752–1761

    CAS  PubMed  Google Scholar 

  21. Sonta T, Inoguchi T, Tsubouchi H, Sekiguchi N, Kobayashi K, Matsumoto S, Utsumi H, Nawata H (2004) Evidence for contribution of vascular NAD(P)H oxidase to increased oxidative stress in animal models of diabetes and obesity. Free Radic Biol Med 37(1):115–123

    Article  CAS  PubMed  Google Scholar 

  22. Erdei N, Tóth A, Pásztor ET, Papp Z, Edes I, Koller A. Bagi Z (2006) High-fat diet-induced reduction in nitric oxide-dependent arteriolar dilation in rats: role of xanthine oxidase-derived superoxide anion. Am J Physiol Heart Circ Physiol 291(5): H2107–H2115

    Article  CAS  PubMed  Google Scholar 

  23. Shinozaki K, Nishio Y, Ayajiki K, Yoshida Y, Masada M, Kashiwagi A, Okamura T (2007) Pitavastatin restores vascular dysfunction in insulin-resistant state by inhibiting NAD(P)H oxidase activity and uncoupled endothelial nitric oxide synthasedependent superoxide production. J Cardiovasc Pharmacol 49(3): 122–130

    Article  CAS  PubMed  Google Scholar 

  24. Moien-Afshari F, Ghosh S, Khazaei M, Kieffer TJ, Brownsey RW, Laher I (2008) Exercise restores endothelial function independently of weight loss or hyperglycaemic status in db/db mice. Diabetologia 51(7):1327–1337

    Article  CAS  PubMed  Google Scholar 

  25. Keenan KP, Laroque P, Dixit R (1998) Need for dietary control by caloric restriction in rodent toxicology and carcinogenicity studies. J Toxicol Environ Health B Crit Rev 1(2):135–148

    CAS  PubMed  Google Scholar 

  26. Ryoo S, Gupta G, Benjo A, Lim HK, Camara A, Sikka G, Lim HK, Sohi J, Santhanam L, Soucy K, Tuday E, Baraban E, Ilies M, Gerstenblith G, Nyhan D, Shoukas A, Christianson DW, Alp NJ, Champion HC, Huso D, Berkowitz DE (2008) Endothelial arginase II: a novel target for the treatment of atherosclerosis. Circ Res 102(8):923–932

    Article  CAS  PubMed  Google Scholar 

  27. Kim F, Pham M, Maloney E, Rizzo NO, Morton GJ, Wisse BE, Kirk EA, Chait A, Schwartz MW (2008) Vascular inflammation, insulin resistance, and reduced nitric oxide production precede the onset of peripheral insulin resistance. Arterioscler Thromb Vasc Biol 28(11):1982–1988

    Article  CAS  PubMed  Google Scholar 

  28. Dikalov S, Griendling KK, Harrison DG (2007) Measurement of reactive oxygen species in cardiovascular studies. Hypertension 49:717–727

    Article  CAS  PubMed  Google Scholar 

  29. Münzel T, Daiber A, Ullrich V, Mülsch A (2005) Vascular consequences of endothelial nitric oxide synthase uncoupling for the activity and expression of the soluble guanylyl cyclase and the cGMP-dependent protein kinase. Arterioscler Thromb Vasc Biol 25(8):1551–1557

    Article  PubMed  Google Scholar 

  30. Palm F, Onozato ML, Luo Z, Wilcox CS (2007) Dimethylarginine dimethylaminohydrolase (DDAH): expression, regulation, and function in the cardiovascular and renal systems. Am J Physiol Heart Circ Physiol 293(6):H3227–H3245

    Article  CAS  PubMed  Google Scholar 

  31. Durante W, Johnson FK, Johnson RA (2007) Arginase: a critical regulator of nitric oxide synthesis and vascular function. Clin Exp Pharmacol Physiol 34(9):906–911

    Article  CAS  PubMed  Google Scholar 

  32. Korda M, Kubant R, Patton S, Malinski T (2008) Leptin-induced endothelial dysfunction in obesity. Am J Physiol Heart Circ Physiol 295(4):H1514–H1521

    Article  CAS  PubMed  Google Scholar 

  33. Siasos G, Tousoulis D, Antoniades C, Stefanadi E, Stefanadis C (2007) l-Arginine, the substrate for NO synthesis: an alternative treatment for premature atherosclerosis? Int J Cardiol 116(3): 300–308

    Article  PubMed  Google Scholar 

  34. McNally JS, Davis ME, Giddens DP, Saha A, Hwang J, Dikalov S. Jo H, Harrison DG (2003) Role of xanthine oxidoreductase and NAD(P)H oxidase in endothelial superoxide production in response to oscillatory shear stress. Am J Physiol Heart Circ Physiol 285(6):H2290–H2297

    CAS  PubMed  Google Scholar 

  35. Landmesser U, Dikalov S, Price SR, McCann L, Fukai T, Holland SM, Mitch WE, Harrison DG (2003) Oxidation of tetrahydrobiopterin leads to uncoupling of endothelial cell nitric oxide synthase in hypertension. J Clin Invest 111(8):1201–1209

    CAS  PubMed  Google Scholar 

  36. Aldieri E, Riganti C, Polimeni M, Gazzano E, Lussiana C, Campia I, Ghigo D (2008) Classical inhibitors of NOX NAD(P)H oxidases are not specific. Curr Drug Metab 9(8):686–696

    Article  CAS  PubMed  Google Scholar 

  37. Das DK, Engelman RM, Clement R, Otani H, Prasad MR, Rao PS (1987) Role of xanthine oxidase inhibitor as free radical scavenger: a novel mechanism of action of allopurinol and oxypurinol in myocardial salvage. Biochem Biophys Res Commun 148(1): 314–319

    Article  CAS  PubMed  Google Scholar 

  38. Symons JD, McMillin SL, Riehle C, Tanner J, Palionyte M, Hillas E, Jones D, Cooksey RC, Birnbaum MJ, McClain DA, Zhang QJ, Gale D, Wilson LJ, Abel ED (2009) Contribution of insulin and Akt1 signaling to endothelial nitric oxide synthase in the regulation of endothelial function and blood pressure. Circ Res 104(9): 1085–1094

    Article  CAS  PubMed  Google Scholar 

  39. Pannirselvam M, Simon V, Verma S, Anderson T, Triggle CR (2003) Chronic oral supplementation with sepiapterin prevents endothelial dysfunction and oxidative stress in small mesenteric arteries from diabetic (db/db) mice. Br J Pharmacol 140(4): 701–706

    Article  CAS  PubMed  Google Scholar 

  40. Minamiyama Y, Bito Y, Takemura S, Takahashi Y, Kodai S, Mizuguchi S, Nishikawa Y, Suehiro S, Okada S (2007) Calorie restriction improves cardiovascular risk factors via reduction of mitochondrial reactive oxygen species in type II diabetic rats. J Pharmacol Exp Ther 320(2):535–543

    Article  CAS  PubMed  Google Scholar 

  41. Ugochukwu NH, Figgers CL (2007) Attenuation of plasma dyslipidemia and oxidative damage by dietary caloric restriction in streptozotocin-induced diabetic rats. Chem Biol Interact 169(1): 32–41

    Article  CAS  PubMed  Google Scholar 

  42. Ungvari Z, Parrado-Fernandez C, Csiszar A, de Cabo R (2008) Mechanisms underlying caloric restriction and lifespan regulation: implications for vascular aging. Circ Res 102(5):519–528

    Article  CAS  PubMed  Google Scholar 

  43. Hoey BM, Butler J, Halliwell B (1988) On the specificity of allopurinol and oxypurinol as inhibitors of xanthine oxidase. A pulse radiolysis determination of rate constants for reaction of allopurinol and oxypurinol with hydroxyl radicals. Free Radic Res Commun 4(4):259–263

    Article  CAS  PubMed  Google Scholar 

  44. Mizia-Stec K, Gasior Z, Zahorska-Markiewicz B, Holecki M, Haberka M, Mizia M, Gomułka S, Zak-Gołab A, Gościńska A (2008) The indexes of arterial structure and function in women with simple obesity: a preliminary study Heart Vessels 23(4): 224–229

    Google Scholar 

  45. Fujii N, Tsuchihashi K, Sasao H, Eguchi M, Miurakami H, Hase M, Higashiura K, Yuda S, Hashimoto A, Miura T, Ura N, Shimamoto K (2008) Insulin resistance functionally limits endothelium-dependent coronary vasodilation in nondiabetic patients Heart Vessels 23(1):9–15

    Google Scholar 

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Authors and Affiliations

  1. Department of Pharmacology and Toxicology, University of Kuopio, Kuopio, Finland

    Juha Ketonen

  2. Institute of Biomedicine, Pharmacology, University of Helsinki, P.O. Box 63, Haartmaninkatu 8, Helsinki, Finland

    Taru Pilvi & Eero Mervaala

  3. Biomedicum Helsinki, University of Helsinki, FIN-00014, Helsinki, Finland

    Taru Pilvi & Eero Mervaala

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  1. Juha Ketonen
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Correspondence to Eero Mervaala.

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Ketonen, J., Pilvi, T. & Mervaala, E. Caloric restriction reverses high-fat diet-induced endothelial dysfunction and vascular superoxide production in C57Bl/6 mice. Heart Vessels 25, 254–262 (2010). https://doi.org/10.1007/s00380-009-1182-x

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  • DOI: https://doi.org/10.1007/s00380-009-1182-x

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